© The Author(s) 2019.In this study, it was aimed to prepare silver nanoparticles by reduction of silver salt (AgNO3) in situ by means of only synthesized polyacrylonitrile (PAN) and poly(acrylonitrile-co-itaconic acid) (P(AN-co-IA)) polymers, and N,N dimethylformamide (DMF). Thereafter, PAN/Ag and P(AN-co-IA)/Ag nanofibers were prepared via electrospinning. Spectroscopic and morphologic characterizations, electrical and thermal features, and antimicrobial activities of the prepared nanofibers against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and Candida albicans were carried out in detail. It was observed that P(AN-co-IA) was much more effective than PAN on the reduction of AgNO3 and formation of silver nanoparticles. Silver nanoparticles and also itaconic acid contributed to decrease the cyclization temperature of PAN by generating sharp exothermic peaks. In addition, electrical conductivity of the nanofibers increased dramatically from E−13 to E−4 related to the presence of silver nanoparticles. Furthermore, incorporation of silver nanoparticles to the nanofiber membranes let bactericidal/fungicidal activities, which started at 6–24 h and continued for up to 168 h, against S. aureus, E. coli, P. aeruginosa, and C. albicans. The prepared silver containing nanofibers can be regarded as good candidates for potential use in the biomedical and pharmaceutical applications.